GFP requires no external substrate for detection, and there have been no reports of detri-
mental effects on the fitness of plants that express it; therefore, it is a nonconditional non-
selectable marker gene. The novelty of GFP and other fluorescent proteins is that they
combine great sensitivity at the subcellular level using bioimaging technologies made avail-
able through confocal laser scanning microscopy with real-time detection in living cells
(Fig. 9.10). A wide variety of fluorescent proteins are being developed to extend the
range and complexity of processes that can be simultaneously monitored in living cells
(Stewart 2006). For example, fluorescent proteins can be fused to various plant proteins
and used as a tag to monitor their trafficking and interactions (Fig. 9.10). In field studies,
GFP also permits the rapid and easy detection of transgenic plants or plant parts such as
pollen (Fig. 9.11). In tissue culture, GFP has been used in combination with selectable
marker genes to identify and enrich the content of transformed material to improve the
recovery of transgenic plants from species where the current transformation and selection
systems are inefficient. New fluorescent proteins are being discovered, and we can expect
a rainbow of colors in the near future (Stewart 2006). Especially useful will be fluorescent
proteins that emit in the orange and red spectra (Fig. 9.12). This is because natural auto-
fluorescence is less in the orange and red spectra in most plants.
9.6 Marker-Free Strategies
Because the roles of the selectable marker genes are often served during the generation of
transgenic plants and not after they have been developed, there is generally no need to
Figure 9.9.Luminescence detected in transgenic tobacco transformed with the firefly luciferase gene
driven by the 35Spromoter and watered with a solution of luciferin, the luciferase substrate.
[Reprinted with permission from Ow et al. (1986), copyright 1986, AAAS.] See color insert.
9.6. MARKER-FREE STRATEGIES 233